Dissolution of Bisphenol A, Bisphenol A Diglycidyl Ether and Its Derivatives in Canned Drinks

金屬罐為主要食品包裝材料之一，一般罐頭內壁會塗佈樹脂，以避免金屬鐵皮腐蝕，但樹脂與食品接觸時，可能會有樹脂原料丙二酚A (bisphenol A, BPA)，或丙二酚A二環氧甘油醚(bisphenol A diglycidyl ether, BADGE)單體溶出。BPA被視為荷爾蒙干擾物質，而BADGE有致癌性疑慮且其化性不穩定，於食品中可能形成衍生物，包括2種水合物[丙二酚A二環氧甘油醚單水合物(bisphenol A (2,3-dihydroxypropyl) glycidyl ether, BADGE•H2O)、丙二酚A二環氧甘油醚二水合物(bisphenol A bis(2,3-dihydroxypropyl) ether, BADGE•2H2O)]及3種含氯化合物[丙二酚A二環氧甘油醚氫氯化物(bisphenol A (3-chloro-2-hydroxypropyl) glycidyl ether, BADGE•HCl)、丙二酚A二環氧甘油醚二氫氯化物(bisphenol A bis(3-chloro-2-hydroxypropyl) ether, BADGE•2HCl)和丙二酚A二環氧甘油醚水合物氫氯化物(bisphenol A (3-chloro-2-hydroxypropyl) (2,3-dihydroxypropyl) ether, BADGE• H2O•HCl)]，亦具有不同程度的毒性。為瞭解金屬罐容器及罐裝飲料中此類化合物可能溶出情形，必須建立一套簡單、快速、穩定之同步檢測法。本研究以高效液相層析法，配合螢光檢出器檢測，使用之層析管柱為Phenomenex Luna C18(2) (25 cm × 4.6 mm i.d., 5 μm thickness)，以乙腈/水/甲醇梯度移動相溶液，於激發波長230 nm、放射波長304 nm可同時檢測上述7種化合物。16件金屬罐檢體採用水、4%醋酸溶液、20%乙醇溶液及正庚烷為食品模擬溶液，進行溶出試驗，結果僅3件檢體檢出BPA，其溶出量為0.002～0.003 mg/dm2；BADGE及其水合物之總溶出量為未檢出～0.065 mg/dm2；含氯化合物之總溶出量為未檢出～0.02 mg/dm2。裝飲料之檢驗方法依基質類型，分別以甲基第三丁基醚或乙腈萃取，經乙腈飽和之正己烷去油脂，再以Sep-Pak C18及Sep-Pak Florisil過濾層析匣淨化，或直接濾膜過濾後，以高效液相層析儀分析定量。添加0.2、0.4及0.8 μg/g混合標準溶液於咖啡檢體中，其平均回收率為80.9～108.8%，變異係數皆小於5.5%；添加0.05、0.1及0.2 μg/g混合標準溶液於番茄汁檢體中，其平均回收率為81.1～109.8%，變異係數皆小於5.7%。本檢驗方法之檢出限量BADGE•2H2O、BADGE•H2O•HCl及BADGE•2HCl均為0.003 ppm，BPA、BADGE、BADGE•H2O及BADGE•HCl均為0.005 ppm。將此方法應用於38件市售罐裝飲料之分析檢驗，結果發現，BPA含量為未檢出～0.173 ppm；BADGE及其水合物之總含量為未檢出～2.695 ppm；含氯化合物之總含量為未檢出～0.663 ppm。以上調查結果均符合歐盟規範。Metal can is a major food packaging material. There is usually a resin coating in the interior wall to protect metal from corrosion. However, the resin in contact with food might result in migration of its components such as bisphenol A (BPA) and the monomer of bisphenol A diglycidyl ether (BADGE). BPA belongs to a group of hor-mone disruptors. BADGE is classified as a mutagen and may readily form hydrolyzed products and chlorohydrin products in food, including bisphenol A (2.3-dihydroxypropyl) glycidyl ether (BADGE•H2O), bisphenol A bis(2,3-dihydroxy propyl) ether (BADGE•2H2O), bisphenol A (3-chloro-2-hydroxypropyl) glycidyl ether (BADGE•HCl), bisphenol A bis(3-chloro-2-hydroxypropyl) ether (BADGE•2HCl), and bisphenol A (3-chloro-2-hydroxypropyl) (2,3-dihydroxypropyl) ether (BADGE• H2O•HCl). These derivatives also have different degrees of toxicity. For monitoring the status of migration of these compounds in metal cans, we need to establish a simple, fast, and stable procedure to analyze these 7 compounds simtanenously. A quantitative method using high performance liquid chromatography (HPLC) coupled with a fluo-rescence detector was therefore developed to assay these compounds. The chroma-tographic separation was accomplished by gradient elution of acetonitrile, water and methanol on a Phenomenex Luna C18(2) column (25 cm × 4.6 mm i.d., 5 μm thickness) with a fluorescence detector at 230 nm excitation and 304 nm emission. Migration tests were performed using water, 4% acetic acid solution, 20% ethanol solution and n-heptane as food simulants in a total of 16 metal cans. The results showed that the mi-gration of BPA occurred in only 3 samples in the range of 0.002～0.003 mg/dm2, and those of BADGE hydrolyzed products and chlorohydrin products were in the range of N.D.～0.065 mg/dm2 and N.D.～0.02 mg/dm2, respectively. epending on the composition of canned drinks, these 7 compounds were ex-tracted with tert-butyl methyl ether or acetonitrile, defatted with n-hexane, cleaned up with Sep-Pak C18 and Florisil, and then analyzed by HPLC. Recovery studies were performed by spiking standard compounds into tomato juice at 0.05, 0.1 and 0.2 μg/g levels and into coffee at 0.2, 0.4 and 0.8 μg/g levels, respectively. Average recoveries in both studies were higher than 80%, and the coefficients of variation were less than 5.7%. The detection limits were 0.003 ppm for BADGE•2H2O, BADGE•H2O•HCl and BADGE•2HCl, and 0.005 ppm for BPA, BADGE, BADGE•H2O and BADGE•HCl. This method was tested in a survey of 38 canned drink samples which were purchased from markets. The results showed that the amounts of bisphenol A, BADGE and its hydro-lyzed and chlorohydrin products were in the range of N.D. ~ 0.173 ppm, N.D. ~ 2.695 ppm, and N.D. ~ 0.663 ppm, respectively, which were in conformity with the regulation of European Union.中文摘要………………………………………………………………….. Ⅰ文摘要………………………………………………………………….. Ⅲ錄……………………………………………………………………….. Ⅴ次……………………………………………………………………….. Ⅶ次……………………………………………………………………….. Ⅷ、前言………………………………………………………………….. 1、研究緣起………………………………………………………….. 1、研究目標………………………………………………………….. 7、文獻整理…………………………………………………………….. 9 一、BPA簡介………………………………………………………….. 9、BADGE及其衍生物……………………………………………… 14 三、環氧樹脂(epoxy resin) …………………………………………… 18 四、傅立葉轉換紅外線光譜………………………………………….. 18 五、溶出試驗………………………………………………………….. 19 六、固相萃取(solid phase extraction, SPE)法………………………… 21 七、衍生化反應……………………………………………………….. 21、材料與方法………………………………………………………….. 24、實驗材料 ………………………………………….……………. 24、化學試藥……………..…………………………………………… 24、設備………………….……………………………………………. 26、實驗方法…………………………………………..……………… 27一)空罐塗料材質鑑別………….…………………….…………… 27二)空罐模擬溶液溶出試驗……………………….….…………… 27三)罐裝飲料試驗溶液之製備…………………………………….. 27四)標準溶液之調製………………………………………………. 29五)HPLC定量分析……………………………………………….. 29六)添加回收試驗…………………………………………………. 31七)再現性及穩定性………………………………….……………. 31八)氣相層析質譜儀確認之操作步驟………………….…………. 31、結果與討論……………………………………………..…………. .. 37、空罐塗料材質鑑別………………………………………..……… 37、高效液相層析條件之探討…………………………….…………. 37一)最適偵測波長之選擇…………………………………………. 37二)移動相之探討……………………………………………….…. 37、標準溶液同日內及異日間之再現性………………………….…. 42、標準曲線………………………………………………………….. 42、空罐溶出試驗…………………………………………….………. 42(一)正庚烷溶出液之分析……………………………….………… 42 (二)BPA及BADGE於溶出條件下安定性………………….…… 48 (三)金屬罐中BPA、BADGE及其衍生物之溶出情形…………… 48 (四)與國外文獻調查結果比較…………………….……………… 59、罐裝飲料之分析………………………………………………….. 59一)本實驗方法之探討……………………………………………. 59二)添加回收試驗及檢出限量…………………………………… 62三)市售罐裝飲料中BPA、BADGE及其衍生物之含量調查……………………………………………………………. 7四)本研究與國外文獻之調查結果………………………………. 67五)健康風險評估………………………………………………… 73六)氣相層析質譜儀確認…………………………………..……. 73、結論……………………………………………………………..…… 75、參考文獻……..……………………………………………………… 77 次 頁次1-1 BADGE及其衍生物官能基……..……………………………… 62-1 罐裝食品中BPA之相關文獻……..……..…………...………… 122-2 罐裝食品中BADGE及其衍生物之相關文獻.………………… 162-3 BPA及BADGE衍生物之三甲基矽化物之分子量…………… 233-1 BPA及BADGE衍生物之三甲基矽化物其特徵離子之質荷比(m/z) …………………………………………………….………. 34-1 BPA、BADGE及其衍生物之同日內及異日間之重複性分析……..……..………………………..……..…………………… 54-2 BPA、BADGE及其衍生物以高效液相層析法分析之線性關係..……..………………………..……..………………………… 74-3 BPA、BADGE及其衍生物的正庚烷溶出液之回收率……..… 494-4 金屬罐體中BADGE及其水合物和含氯化合物於不同溶出試驗之溶出情形……………………..……..……………………… 84-5 本研究及文獻之BPA溶出試驗數據……………...…………… 604-6 咖啡飲料中添加BPA、BADGE及其衍生物之回收率.….…… 684-7 番茄汁中添加BPA、BADGE及其衍生物之回收率.………… 694-8 罐裝飲料中BPA、BADGE與其水合物及含氯化合物之含量……..……..…………………………..……..………………… 14-9 罐裝飲料中BPA含量之調查結果..……..………………..…… 72 次 頁次1-1 環氧樹脂分子中所含環氧基(環氧乙烷) …..………..……... 31-2 BADGE合成..……..…………………..……..………………... 31-3 BPA型之聚合物……………………..……..………………..... 41-4 BADGE•HCl衍生物之形成…………..……..………………... 41-5 BADGE衍生物之形成………………..……..………………... 52-1 BPA於聚碳酸酯(PC)、環氧樹脂(ER)及其他產品之全球年度需求量………………………………………………………. 02-2 傅立葉轉換紅外線光譜分析儀操作原理示意圖..…………... 202-3 BSTFA + TMCS矽烷化反應機制..………………………....... 223-1 BPA三甲基矽化物及BADGE之質譜圖…………….………. 343-2 BADGE•HCl及BADGE•H2O三甲基矽化物之質譜圖…….... 353-3 BADGE•2HCl、BADGE•H2O•HCl及BADGE•2H2O三甲基矽化物之質譜圖……................................................................. 64-1 環氧樹脂之紅外光圖譜………..……..………………............. 384-2 聚酯之紅外光圖譜..………………... ..………………... ..…... 394-3 聚丙烯酸及聚甲基丙烯酸酯之紅外光圖譜………..……..…. 404-4 BPA及BADGE之螢光圖譜………..……….…..……………. 414-5 移動相溶液不同甲醇濃度對於BPA等化合物分離之影響… 434-6 BPA、BADGE及其衍生物標準品之HPLC圖譜…….…….. 444-7 BPA、BADGE及其衍生物之標準曲線.. ……………..……... 464-8 BPA於不同溶出試驗之安定性.. ……………..……................ 504-9 BADGE於不同溶出試驗之安定性.. ………..…….................. 514-10 金屬罐體中BPA、BADGE與其水合物及含氯化合物於水(60℃，30分鐘)之溶出情形………..……............................... 24-11 金屬罐體中BPA、BADGE與其水合物及含氯化合物於水(95℃，30分鐘)之溶出情形………..…………........................ 34-12 金屬罐體中BPA、BADGE與其水合物及含氯化合物於4%醋酸溶液(60℃，30分鐘)之溶出情形………..…………........ 44-13 金屬罐體中BPA、BADGE與其水合物及含氯化合物於4%醋酸溶液(95℃，30分鐘)之溶出情形……………...……....... 54-14 金屬罐體中BPA、BADGE與其水合物及含氯化合物於20%乙醇溶液(60℃，30分鐘)之溶出情形………….…..……...... 64-15 金屬罐體中BPA、BADGE與其水合物及含氯化合物於正庚烷(25℃，1小時)之溶出情形………..…………….................. 74-16 不同沖提溶液對於BPA、BADGE及其衍生物自Oasis HLB層析匣溶離回收率之影響………..……….….......................... 34-17 二種不同沖提溶液對於BPA、BADGE及其衍生物自Oasis HLB層析匣溶離回收率之比較….………….......................... 44-18 以甲醇/乙腈(2/8, v/v)沖提液自Sep-pak C18層析匣溶離BPA、BADGE及其衍生物之效果…………...…….................. 54-19 不同比例乙腈/水流洗液自Sep-pak C18層析匣溶離BPA、BADGE及其衍生物效果之比較………..……..….................. 64-20 罐裝咖啡飲料及番茄汁檢體中添加BPA、BADGE及其衍生物之層析圖譜………..………………....................................... 04-21 BPA、BADGE及其衍生物標準品及咖啡檢體之氣相質譜層析圖………………………………………………………..…...